Telomerase activity during spontaneous immortalization of Li-Fraumeni syndrome skin fibroblasts

Abstract

Li-Fraumeni Syndrome (LFS) is characterized by heterozygous germline mutations in the p53 gene. Accompanied by genomic instability and loss or mutation of the remaining wild type p53 allele, a low frequency of spontaneous immortalization in LFS ®broblasts occurs. It is believed that the loss of p53 wild type function contributes to immortalization of these LFS ®broblasts, but it is not clear if this is su cient. Because stabilization of telomere length is also thought to be a necessary step in immortalization, telomerase activity, expression of the telomerase RNA component (hTR) and telomere length were analysed at various passages during the spontaneous immortalization of LFS skin ®broblasts. One LFS strain which immortalized, MDAH087 (087), had no detectable telomerase activity whereas another LFS strain which immortalized, MDAH041 (041), had detectable telomerase activity. In preimmortal cells from both strains, hTR was not detected by in situ hybridization. Immortal 087 cells remained negative for hTR, while immortal 041 cells demonstrated strong hTR in situ hybridization signals. 087 cells had long and heterogenous telomeres whereas telomeres of 041 cells had short, stable telomere lengths. Tumorigenicity studies in nude mice with ras-transformed 087 and 041 cells resulted in both cell lines giving rise to tumors and retaining telomerase status. Overall these results suggest that strain speci®city may be important in telomerase re-activation and that both abrogation of p53 function and a mechanism to maintain telomeres are necessary for immortalization.

Human fibroblasts require the Rb family of tumor suppressors, but not p53, for PML-induced senescence

“…Total proteins (20 mg) were used for SDS-PAGE and individual proteins were visualized after immunoblotting using specific antibodies senescence, indicating that in human fibroblasts, the p53 pathway is not essential for PML-induced senescence. Given the extensive literature implicating p53 in the senescent response of fibroblasts (Gollahon et al, 1998;Campisi, 2001), our results were surprising. However, p53 was reported to be nonessential for the senescent response of human fibroblasts to oncogenic ras (Serrano et al, 1997) or telomere deprotection (Smogorzewska and de Lange, 2002).…”

Epigenetic silencing of multiple interferon pathway genes after cellular immortalization

“…MDAH041 LFS cells contain significant telomerase activity after immortalization (Gollahon et al, 1998). Although in microarray analysis, the hTERT gene for the protein of enzymatic subunit of telomerase was not significantly upregulated after immortalization of MDAH041 cells, 1.6-fold, we found using Q-RT-PCR that there was a significant increase in hTERT expression, 486-fold (Table 2).…”

Combination therapy of malignant glioma cells with 2-5A-antisense telomerase RNA and recombinant adenovirus p53

“…46 Additionally, telomerase activity was unaffected by overexpression of p53 in immortalized endothelial cells. 47 Conversely, it was reported that induced expression of wild-type p53 resulted in the down-regulation of telomerase activity in not only immortalized fibroblasts, 48 but also cancer cell lines. 49,50 Furthermore, it has recently been demonstrated that severe telomere shortening in telomerase-deficient mice activated p53, leading to growth arrest and/or apoptosis.…”